1. Technical Field of the Invention
The invention relates to a mixture of natural microorganisms, which, in its special composition, is capable of biologically degrading hydrocarbons. The invention is especially useful in degrading mineral oil, mineral oil by-products and related hydrocarbons. Further, the invention relates to the use of this mixture for the biodegradation of hydrocarbons. The invention teaches a special process for the cultivation and application of the mixture for the biodegradation of oil-contaminated soils and waters.
2. Description of the Prior Art
The interest in the focussed use of microorganisms for the biological degradation of mineral oil, its derivates and products has increased in the recent years. As early as in 1903, the first organisms have been described which use hydrocarbons as a source for carbons and energy. At present, more than 40 species of microorganisms are known which use aliphatic or aromatic hydrocarbons in the course of a biological degradation. These organisms have been isolated from the oceans, from sweet-waters as well as from the soil.
The number of bacteria and mushrooms which are able to biologically degrade mineral-oil hydrocarbons, increases sharply and locally after oil spills. These substances occur in a substantial part as selection-gas.
From a genetic view point it is clear that the ability of a major proportion of microorganisms to degrade mineral-oil hydrocarbons is controlled by plasmid. By the proof of the transfer of plasmid it became possible to explain the sharp increase of the number of organisms with degrading ability.
As a subtrate for microorganisms, mineral oil is an extremely complex mixture of hydrocarbons. The saturated fraction of mineral oil includes n-alkanes, branching alkanes and cyclo-alkanes. The biodegradation of the n-alkanes up to C.sub.44 is in most cases realized by the monoterminal attack under the creation of carbonacids. This process may lead to an accumulation of some more or less toxic fat-acids.
The branching isoprenoid alkanes are degraded by w-oxidation under the creation of dicarbon-acids. The terminal branched alkanes are very resistant to a biodegradation. They block the .beta.-oxidation and have the tendency to accumulate within the milieu. After the spontaneous degradation of a hydrocarbon-contamination, tripentacyclic bindings are often created, which are persistent in the milieu. Because of that it is clear that the qualitative composition of the oil and its products have a high influence on their degradability.
In the metabolism of the mineral-oil hydrocarbons two other processes play an important role--the cooxidation and prolongation. Many oil-components which are not degradable according to the opinion of many skilled professionals, are degradable when they occur in presence of other hydrocarbons which allow the growth of the microorganisms. The degradation process of microorganisms by the inventive mixture is in no way comparable with the ability of the pure species to degrade the single hydrocarbons.
The speed of the microbial degradation of the mineral-oil hydrocarbons is influenced by many factors which directly or indirectly have an influence on the growth and the metabolism of the microbial population. As crucial appears the physical condition of the hydrocarbons, since the hydrocarbons are degraded the most intensively on the "separation-line" between water and hydrocarbon. Further abiotic factors are the temperature and the nutrions which are available. Most heterotroph microorganisms are able to degrade hydrocarbons within a wide temperature range of 0.degree. C. to 70.degree. C. The temperature not only has an influence on the speed of the degradation but above all on the physical condition of the hydrocarbons, which is in a close connection to the selective pressure of the temperature on microbial communities which are created by the eco-system. In the course of the complex evaluation, the temperature must be considered in connection with other factors. In any case, the temperature must not necessarily be a limiting factor.
The speed of the degradation of mineral oil-products is substantially influenced by the presence of nitrogen and phosphor, which are commonly incorporated in the biomass with hydrocarbon-containing splitting substances. The optimal mass-ratios between hydrocarbon and nitrogen on the one side and between hydrocarbon and phosphor on the other side depend on the qualitative composition of the mineral oil or the mineral oil-products and of the abiotic factors of the milieu. Nitrogen and phosphor are also the common components of "stimulators" which stimulate the biodegradation processes.
For the industrial application of microorganisms in the course of the reclamation of contaminated soils or waters, the degradation rate and the quality of the degradation are of highest interest. For the acceleration of the degradation rate, the following distinct main-categories are known:
1. Fertilization: This method is restricted to only the optimalization of the milieu for the microorganisms which are inherent in the contaminated soil or water. One takes it as a basis that in such a contaminated soil or water, oil-degrading microorganisms in sufficient quality and quantity are present anyway, and concentrates the activities to the addition of nitrogen, phosphor, calcium as well as some trace elements, and to the support of the breathing ability of the present, always aerobic microorganisms by supplying air. For this purpose, the soil is loosened and turned on a regular basis, or air is being pumped into the soil by means of air-lines and pumps. In many cases, a heap of several meters height is equipped with a number of perforated tubes going through the heap on several levels whereby the tubes are being put under air-pressure or vacuum (vacuum-heap-system). The professionals favor this system and are confident that it will be most successful since one thinks that after an oil spill, oil-degrading microorganisms will quickly be present in sufficient quantity, and the degradation is only restricted by the lack of nutrition- and air-supply for the organisms. PA1 2. Seeding: As a separate technique or additional to the above mentioned method, the seeding-technique is used. Seeding means the application of additional oil-degrading microorganisms into the contaminated material. Such preparations which are used by different companies on sites with oil-contaminations usually contain only a single species. The application of such preparations is adequate for fresh contaminations where eventual products of the metabolism are present which appear because of the natural present organisms. The opinion is established among the professionals that the additional application of more natural microorganisms does not substantially increase the effectiveness of the degradation. Moreover, the public in general has some reservations against the transplantation of foreign microorganisms since--be it reasonable or not--one is afraid of there uncontrolled growth, accompanied by unnecessary side effects, which may even be toxic. PA1 3. GEM: This is an abbrevation for "Genetically Engineered Microorganisms", which have very effective qualities in view of their oil-degradation capability. The reservations against the use of such GEM's are the strongest. Therefore, their application will most likely be more and more restricted.